gp96 (GRP94) elicits antigen-presenting cell (APC) activation and can direct peptides into the cross- presentation pathways of APC. These responses arise through interactions of gp96 with Toll-like (APC activation) and endocytic (cross-presentation) receptors of APC. Previously, CD91, the alpha2-macroglobulin receptor, was identified as the heat shock/chaperone protein receptor of APC. Recent data indicates, however, that inhibition of CD91 ligand binding does not alter gp96 recognition and uptake. Furthermore, CD91 expression is not itself sufficient for gp96 binding and internalization. We now report that scavenger receptor class-A (SR-A), a prominent scavenger receptor of macrophages and dendritic cells, serves a primary role in gp96 and calreticulin recognition and internalization. gp96 internalization and peptide re-presentation are inhibited by the SR-A inhibitory ligand fucoidin, although fucoidin was without effect on alpha2-macroglobulin binding or uptake. Ectopic expression of SR-A in HEK 293 cells yielded gp96 recognition and uptake activity. In addition, macrophages derived from SR-A-/- mice were substantially impaired in gp96 binding and uptake. These data identify new roles for SR-A in the regulation of cellular responses to heat shock proteins.

During infection, signals from the periphery are known to reach draining lymph nodes (DLNs), but how these molecules, such as inflammatory cytokines, traverse the significant distances involved without dilution or degradation remains unclear. We show that peripheral mast cells, upon activation, release stable submicrometer heparin-based particles containing tumor necrosis factor and other proteins. These complexes enter lymphatic vessels and rapidly traffic to the DLNs. This physiological drug delivery system facilitates communication between peripheral sites of inflammation and remote secondary lymphoid tissues.

Pseudomonas aeruginosa is a pathogenic Gram-negative bacterium that causes severe opportunistic infections in immunocompromised individuals; in particular, severity of infection with P. aeruginosa positively correlates with poor prognosis in cystic fibrosis (CF) patients. Establishment of chronic infection by this pathogen is associated with downregulation of flagellar expression and of other genes that regulate P. aeruginosa motility. The current paradigm is that loss of flagellar expression enables immune evasion by the bacteria due to loss of engagement by phagocytic receptors that recognize flagellar components and loss of immune activation through flagellin-mediated Toll-like receptor (TLR) signaling. In this work, we employ bacterial and mammalian genetic approaches to demonstrate that loss of motility, not the loss of the flagellum per se, is the critical factor in the development of resistance to phagocytosis by P. aeruginosa. We demonstrate that isogenic P. aeruginosa mutants deficient in flagellar function, but retaining an intact flagellum, are highly resistant to phagocytosis by both murine and human phagocytic cells at levels comparable to those of flagellum-deficient mutants. Furthermore, we show that loss of MyD88 signaling in murine phagocytes does not recapitulate the phagocytic deficit observed for either flagellum-deficient or motility-deficient P. aeruginosa mutants. Our data demonstrate that loss of bacterial motility confers a dramatic resistance to phagocytosis that is independent of both flagellar expression and TLR signaling. These findings provide an explanation for the well-documented observation of nonmotility in clinical P. aeruginosa isolates and for how this phenotype confers upon the bacteria an advantage in the context of immune evasion.Pseudomonas aeruginosa is an opportunistic Gram-negative bacterial pathogen that causes severe infections in immunocompromised patients and in the pulmonary compartment of patients suffering from cystic fibrosis (CF) (13,14). In CF patients, disease severity is positively correlated with colonization by P. aeruginosa and the establishment of chronic infection. As part of the colonization process, the bacteria undergo a number of genetic changes that assist in their ability to survive in the mammalian host and to evade detection and clearance by the immune system (9, 21). One such change that has been phenotypically characterized for P. aeruginosa is loss of flagellar motility (12,17). Furthermore, the loss of flagellar gene expression and motility function is associated with increased bacterial burdens and increased disease severity in CF patients (12,17). While downregulation of flagellar expression has been inferred to confer a survival advantage on P. aeruginosa once it colonizes the host by evasion of both phagocytic receptors and TLR5-driven inflammatory signaling, the exact contribution of flagellum downregulation with respect to successful immune evasion is unclear (5, 17, 18).Nonopsonic phagocytosis of P. aeruginosa by murine and human mac...

CAPS is a neural/endocrine-specific protein discovered as a cytosolic factor required for Ca2+-activated dense-core vesicle (DCV) exocytosis in permeable neuroendocrine cells. We report that CAPS is also a membrane-associated, peripherally bound protein in brain homogenates that localizes Selectively to plasma membranes and to DCVs but not to small clear synaptic vesicles (SVs). CAPS exhibits high affinity and saturable binding to DCVs by interaction with bilayer phospholipids. Specific CAPS antibodies inhibit Ca2+-activated norepinephrine release from lysed synaptosomes that contain membrane-associated CAPS, indicating that membrane-bound CAPS is essential for neural DCV exocytosis. CAPS is a functional component of the exocytotic machinery that localizes selectively to DCVs, and it may confer distinct regulatory features on neuropeptide and biogenic amine transmitter secretion.